MicroRNAs (miRNAs) are short non-coding RNAs that control gene expression predominantly through post- transcriptional repression, implicated in the control of multiple physiological and pathological processes. Recently, we described roles for endothelial miRNAs in central aspects of vascular biology such as angiogenesis and inflammation, processes that require the coordination of numerous and complex signaling pathways in which endothelial cells (ECs) act as both active participants and regulators. Endothelial activation is essential for many vascular processes leading to inflammation, vascular remodeling and vessel growth. Cytokines, such as vascular endothelial growth factor (VEGF) and tumor necrosis factor (TNF) mediate this activation and play fundamental roles in the control of angiogenic and inflammatory responses, respectively. miRNA function is modulated by extracellular factors affecting their biogenesis or activity, thereby fine-tuning the regulation of biological responses. However, our understanding of the mechanisms governing the regulation of microRNA in ECs is limited. We postulate that physiological levels of miRNAs are regulated under conditions, such as inflammation or angiogenic activation, to coordinate specific gene expression programs in ECs. We propose three aims: Aim 1: To identify the molecular mechanisms whereby VEGF and TNF regulate miRNA levels in ECs. Aim 2: To examine the regulation of EC targets and functions by VEGF and TNF-regulated miRNA in vitro. Aim 3: To define the roles of VEGF and TNF- regulated miRNAs in vivo. In summary, completion of these aims will provide critical insight into fundamental regulatory mechanisms controlling endothelial miRNA biogenesis and activity and their impact on EC biology. Understanding the complex network involving EC miRNAs and their targets, leading to a coordinate pattern of gene expression undoubtedly will contribute to a better understanding of the cellular and molecular mechanisms that control EC activation and may identify potential therapeutic strategies for the regulation of endothelial activation through modulation of miRNA activity! PUBLIC HEALTH RELEVANCE: We have described roles for endothelial microRNAs in central aspects of vascular biology such as angiogenesis and inflammation. The proposal main goal is to investigate the mechanisms that regulate microRNA biogenesis and functions in endothelial cell activation. This work will provide critical insight into fundamental regulatory mechanisms and may identify potential therapeutic strategies for the regulation of endothelial activation through modulation of microRNA activity.